Project Summary Visual clarity depends on strict vascular demarcations between ocular vascular beds and vision-critical tissues such as the cornea and macular photoreceptors. Our long-range goals are to elucidate the molecular network that maintains this compartmentalization in the eye and to understand how the network is altered in pathological ocular angiogenesis. Our immediate goal is to dissect tissue-specific differences in vascular versus neuronal requirements for vascular endothelial growth factor (VEGF) under normal and pathological conditions. We will determine how soluble VEGF receptors orchestrate cross-talk among corneal compartments to titrate the balance between VEGF requirements for vascular demarcation and neuronal homeostasis. We will genetically dissect the roles of specific ocular compartments to determine how VEGF-dependent nerve regeneration is accomplished without triggering a vascular breach. We will rigorously analyze the vascular response of soluble VEGFR1 (sFlt1) knockdown within each corneal compartment during epithelial wound healing and nerve regeneration. Complementing this analysis, we will dissect the roles of sVEGF receptors expressed on corneal neurons. Finally, we will elucidate whether splice shifting morpholinos conjugated to a neovessel-targeting peptide motif can treat corneal neovascularization (KNV) and choroidal neovascularization (CNV). Our specific aims for the next grant period are: Specific Aim #1: To rigorously test functional requirements for soluble VEGFR1 (sFlt1) or soluble VEGFR2 (sKDR) within specific corneal compartments for achieving vascular demarcation and appropriate innervation in the normal and injured cornea. Specific Aim #2: To determine whether aniridia-associated KNV resulting from Pax6 haploinsufficiency derives from neural crest related defects in the corneal stroma that shift angiogenic potential, and whether vascular breach from limbal stem cell deficiency is a secondary paracrine mechanism due to epithelial upregulation of MT2-MMP. Specific Aim #3: To determine whether local administration of RGD-conjugated splice shifting morpholino (cRGD.KDR.MO) can significantly increase sKDR/mKDR ratios and decrease neovascularization in laser-CNV injury and sutured corneas of wild type mice, and in Pax6+/- mouse corneas. These studies will define the tissue-specific functions of sFlt1 and sKDR in the context of competing requirements for neuronal and vascular homeostasis in the cornea, systematically detailing the development and maintenance of vascular zoning and neurite outgrowth in normal and pathological situations.